Expression of the MDR1 and MRP genes in patients with lymphoma with primary bone marrow involvement

Expression of MDR1 and MRP genes in patients with low-grade and high-grade non-Hodgkin's lymphomas with primary bone marrow involvement before and after chemotherapy was investigated. The data demonstrate that overexpression of MDR1 and MRP genes in hematological malignancies elevates in patients after chemotherapy and correlates with poor clinic prognosis and more frequent recurrences of the malignancies.     

miR‐495 sensitizes MDR cancer cells to the combination of doxorubicin and taxol by inhibiting MDR1 expression

MDR1 is highly expressed in MDR A2780DX5 ovarian cancer cells, MDR SGC7901R gastric cancer cells and recurrent tumours. It pumps cytoplasmic agents out of cells, leading to decreased drug accumulation in cells and making cancer cells susceptible to multidrug resistance. Here, we identified that miR‐495 was predicted to target ABCB1, which encodes protein MDR1. To reduce the drug efflux and reverse MDR in cancer cells, we overexpressed a miR‐495 mimic in SGC7901R and A2780DX cells and in transplanted MDR ovarian tumours in vivo. The results indicated that the expression of MDR1 in the above cells or tumours was suppressed and that subsequently the drug accumulation in the MDR cells was decreased, cell death was increased, and tumour growth was inhibited after treatment with taxol‐doxorubicin, demonstrating increased drug sensitivity. This study suggests that pre‐treatment with miR‐495 before chemotherapy could improve the curative effect on MDR1‐based MDR cancer.     

Drug resistance and IS6110‐RFLP patterns of Mycobacterium tuberculosis in patients with recurrent tuberculosis in northern Thailand

The emergence of drug resistant Mycobacterium tuberculosis has become a global threat to tuberculosis (TB) prevention and control efforts. This study aimed to determine the drug resistance profiles and DNA fingerprints of M. tuberculosis strains isolated from patients with relapsed or retreatment pulmonary TB in Chiang Rai province in northern Thailand. Significant differences in multidrug resistance (MDR) (P = 0.025) and resistance to isoniazid (P = 0.025) and rifampin (P = 0.046) between first and second registrations of patients with retreatment TB were found. However, there were no significant differences in resistance to any drugs in patients with relapsed TB. The rate of MDR‐TB strains was 12.2% among new patients at first registration, 22.5% among patients with recurrence who had previously undergone treatment at second registration and 12.5% at third registration. Two retreatment patients whose initial treatment had failed had developed MDR‐TB with resistance to all TB drugs tested, including rifampin, isoniazid, streptomycin and ethambutol. IS6110‐RFLP analysis revealed that 66.7% (10/15 isolates) of MDR‐TB belonged to the Beijing family. In most cases, IS6110‐RFLP patterns of isolates from the same patients were identical in relapse and retreatment groups. However, some pairs of isolates from retreatment patients after treatment failure had non‐identical IS6110‐RFLP patterns. These results suggest that, after failure and default treatment, patients with retreatment tuberculosis have a significantly greater risk of MDR‐TB, isoniazid and rifampin resistance than do other patients.     

ARA,a Novel ABC Transporter, Is Located at 16p13.1, Is Deleted in inv(16) Leukemias, and Is Shown To Be Expressed in Primitive Hematopoietic Precursors

ATP-binding cassette (ABC), ATP-dependent transporters are a large superfamily of proteins that include the multidrug resistance proteins, P-glycoprotein and MRP (multidrug resistance protein). TheARA(anthracycline resistance-associated) gene that codes for a putative member of the ABC transporters has recently been cloned and shown to have high sequence homology to the gene for MRP. We have previously shownMRPto be deleted in a subset of inv(16) leukemic patients. The deletion ofMRPwas associated with an improved patient survival compared with inv(16) patients who did not have such a deletion. In this study, theARAgene is mapped to 16p13.1, in the same physical interval as the inv(16) short-arm breakpoint. It is shown to be situated proximal to bothMYH11,the gene involved in the primary breakpoint on the short arm of the inv(16), andMRP.A YAC clone has been isolated containing bothMRPandARA.FISH analysis of metaphase chromosomes from inv(16) patients has established the gene order as telomere–MYH11–MRP–ARA–centromere and demonstrated that bothARAandMRPare deleted in a subgroup of the inv(16) leukemias.ARAandMRPare both shown to be expressed in normal hematopoietic precursors including CD34⁺cells. The mapping ofARAto this region and its homology toMRPraises questions about its potential role in the biology of the inv(16) leukemias.     

Calpain-2 expression is associated with response to platinum based chemotherapy, progression-free and overall survival in ovarian cancer

Ovarian cancer is routinely treated with surgery and platinum‐based chemotherapy. Resistance is a major obstacle in the efficacy of this chemotherapy regimen and the ability to identify those patients at risk of developing resistance is of considerable clinical importance. The expression of calpain‐1, calpain‐2 and calpastatin were determined using standard immunohistochemistry on a tissue microarray of 154 primary ovarian carcinomas from patients subsequently treated with platinum‐based adjuvant chemotherapy. High levels of calpain‐2 expression was significantly associated with platinum resistant tumours (P = 0.031). Furthermore, high expression of calpain‐2 was significantly associated with progression‐free (P = 0.049) and overall survival (P = 0.006) in this cohort. The association between calpain‐2 expression and overall survival remained significant in multivariate analysis accounting for tumour grade, stage, optimal debulking and platinum sensitivity (hazard ratio = 2.174; 95% confidence interval = 1.144–4.130; P = 0.018). The results suggest that determining calpain‐2 expression in ovarian carcinomas may allow prognostic stratification of patients treated with surgery and platinum‐based chemotherapy. The findings of this study warrant validation in a larger clinical cohort.     

Lack of association between <Emphasis Type="Italic">MDR1</Emphasis> C3435T polymorphism and chemotherapy response in advanced breast cancer patients: evidence from current studies

The transmembrane transport of anticancer drugs is mainly regulated by P-glycoprotein encoded by the human multidrug resistance gene 1 gene (MDR1). Since there were controversies regarding the association between MDR1 C3435T polymorphism and response to chemotherapy among patients with advanced breast cancer, a meta-analysis of the link was conducted. A total of 7 studies consist of 464 advanced breast cancer patients relating MDR1 C3435T polymorphism to the response of chemotherapy were included in this meta-analysis. The main analysis revealed a lack of association between the MDR1 C3435T and response to chemotherapy, with odds ratios (ORs) and their corresponding 95% confidence intervals (CIs) of 1.37 (95% CI: 0.78–2.40), 1.17 (95% CI: 0.69–2.01), 1.18 (95% CI: 0.76–1.84) and 1.61 (95% CI: 0.70–3.68) for homozygous comparison, heterozygous comparison, dominant model and recessive model, respectively. The subgroup analysis by ethnicity did not change the pattern of results, with ORs of 0.99 (95% CI: 0.11–9.07), 0.68 (95% CI: 0.29–1.60), 0.81 (95% CI: 0.36–1.85) and 1.51 (95% CI: 0.77–2.96), in homozygous comparison, heterozygous comparison, dominant model and recessive model, respectively in Caucasian, and 1.50 (95% CI: 0.75–3.03), 1.72 (95% CI: 0.85–3.47), 1.59 (95% CI: 0.90–2.80) and 2.29 (95% CI: 0.51–10.35), respectively in Asian. The available evidence indicates that MDR1 C3435T polymorphism cannot be considered as a reliable predictor of response to chemotherapy in patients with advanced breast cancer.     

The influence of proteasome inhibitor bortezamib on ABC transporters’ expression and activity in tumor cells

The goal of this study was to investigate the role of the ABC transporters in the evolution of tumor cell populations treated with bortezomib. Bortezomib (PS-341, Velcade) is a proteasome inhibitor used for treatment of some malignancies. Several pairs of cell lines different in Pgp expression (P-glycoprotein transporter, ABCB1) have been used in the study. We showed that the influence of the Pgp hyperexpression on cell sensitivity to bortezomib was bidirectional and depended on the tissue type. Bortezomib changed the mRNA level of MDR1 (ABCB1 and MRP1 (ABCC1)) genes, suggesting that the proteasome inhibitor is able to decrease the activity of some regulators of genes/proteins implicated in MDR. Bortezomib treatment increased the levels of proteins (Pgp or MPR1) in 3 out of 4 cell populations studied. Pgp was shown to remain functionally active in the cells cultured in bortezamib-containing medium. The UIC2-shift assay has shown that bortezomib is able to activate Pgp. This means that bortezomib influences Pgp conformation, thus activating the protein (in K562/i-S9 cells). These experiments also demonstrate that bortezomib is Pgp substrate.     

EFFECTS OF GENETIC KNOCK‐DOWN OF ORGANIC ANION TRANSPORTER GENES ON SECRETION OF FLUORESCENT ORGANIC IONS BY MALPIGHIAN TUBULES OF Drosophila melanogaster

An earlier study has shown that RNAi knock‐down of a single organic anion transporter (OAT) gene in the principal cells of Drosophila Malpighian tubules is associated with reductions in the expression of multiple, functionally related genes. In this study, we measured the rates of secretion of four fluorescent ions by tubules isolated from flies expressing targeted RNAi knock‐down of specific OAT genes. Droplets secreted by isolated tubules set up in the Ramsay assay were collected in optically flat capillary tubes and the concentrations of fluorescent ions were determined by confocal laser scanning microscopy. Reductions in the expression of organic anion (OA) transporting polypeptide 58Dc (OATP; CG3380) were associated with reduced secretion of the OAs fluorescein and Texas Red. Reduction in the expression of Drosophila multidrug resistance associated protein (dMRP; CG6214) was correlated with reduced secretion of the P‐glycoprotein substrate daunorubicin. Secretion of the organic cation quinacrine was unaffected by reduced expression of OATP, dMRP, or a multidrug efflux transporter (MET; CG30344). The results highlight the difficulties of assigning a rate‐limiting role in transport of a specific OA to a single membrane transporter.     

Fungicide efflux and the MgMFS1 transporter contribute to the multidrug resistance phenotype in Zymoseptoria tritici field isolates

Septoria leaf blotch is mainly controlled by fungicides. Zymoseptoria tritici, which is responsible for this disease, displays strong adaptive capacity to fungicide challenge. It developed resistance to most fungicides due to target site modifications. Recently, isolated strains showed cross‐resistance to fungicides with unrelated modes of action, suggesting a resistance mechanism known as multidrug resistance (MDR). We show enhanced prochloraz efflux, sensitive to the modulators amitryptiline and chlorpromazine, for two Z. tritici strains, displaying an MDR phenotype in addition to the genotypes CYP51^{I381V Y461H} or CYP51^{I381V ΔY459/G460}, respectively, hereafter named MDR6 and MDR7. Efflux was also inhibited by verapamil in the MDR7 strain. RNA sequencing lead to the identification of several transporter genes overexpressed in both MDR strains. The expression of the MgMFS1 gene was the strongest and constitutively high in MDR field strains. Its inactivation in the MDR6 strain abolished resistance to fungicides with different modes of action supporting its involvement in MDR in Z. tritici. A 519 bp insert in the MgMFS1 promoter was detected in half of the tested MDR field strains, but absent from sensitive field strains, suggesting that the insert is correlated with the observed MDR phenotype. Besides MgMfs1, other transporters and mutations may be involved in MDR in Z. tritici.     

Modulation of Multidrug Resistance in Cancer Cells by Liposome Encapsulated Doxorubicin

Abstract

A major problem in the chemotherapy of solid tumors and hematologic malignancies is the intrinsic as well as acquired cross resistance to multiple chemotherapeutic agents. Recently, this type of multidrug resistance has been related to a gene, MDR1, and its gene product, p-glycoprotein, which functions as the efflux pump, prevents accumulation of drugs and alters their cytotoxicity. Many drug-resistant human tumors express the MDR1 gene and MDR1 RNA levels are elevated in many cancers that have not responded to chemotherapy. The same persistent observation has been made in recurrent tumors who have responded initially to chemotherapy.

Doxorubicin is one of the most important anticancer agent having significant single agent activity in a variety of cancer types and is now the cornerstone of some widely used combination regimens. Despite the clinical effectiveness of the drug, doxorubicin resistance that arises in malignant cells following repeated courses of treatment is the major problem in the clinical management of neoplastic diseases. Recently, extensive studies have demonstrated that liposome encapsulated doxorubicin effectively modulates the multidrug resistance phenotype in cancer cells by altering the function of p-glycoprotein. This modulation of MDR phenotype by liposomes has been demonstrated in vitro in human breast cancer cells, ovarian cancer cells, human promyelocytic leukemia cells and in human colon cancer cells and in vivo in transgenic mice transfected with a functional MDR1 gene. It appears liposomes can play an effective role as a new modality of treatment for human cancers which have become refractory to chemotherapy. An exciting area of research which soon will emerge will exploit the different binding sites on p-glycoprotein by using combination of liposomes with other pharmacological modulators of MDR to impart maximal overcoming of multidrug resistance in cancer patients.     

Association of NRG1 and AUTS2 genetic polymorphisms with Hirschsprung disease in a South Chinese population

Hirschsprung disease (HSCR) is a genetic disorder characterized by the absence of enteric ganglia. There are more than 15 genes identified as contributed to HSCR by family‐based or population‐based approaches. However, these findings were not fulfilled to explain the heritability of most sporadic cases. In this study, using 1470 HSCR and 1473 control subjects in South Chinese population, we replicated two variants in NRG1 (rs16879552, P = 1.05E‐04 and rs7835688, P = 1.19E‐07), and further clarified the two replicated SNPs were more essential for patients with short‐segment aganglionosis (SHSCR) (P = 2.37E‐05). We also tried to replicate the most prominent signal (rs7785360) in AUTS2, which was a potential susceptibility gene with HSCR. In our results, in terms of individual association, marginal effect was observed to affect the HSCR patients following recessive model (P = 0.089). Noteworthy, significant intergenic synergistic effect between rs16879552 (NRG1) and rs7785360 (AUTS2) was identified through cross‐validation by logistic regression (P = 2.45E‐03, OR = 1.53) and multifactor dimensionality reduction (MDR, P < 0.0001, OR = 1.77). Significant correlation was observed between expression of these two genes in the normal segments of the colons (P = 0.018), together with differential expression of these genes between aganglionic colonic segments and normal colonic segments of the HSCR patients (P value for AUTS2 <0.0001, P value for NRG1 = 0.0243). Although functional evaluation is required, we supply new evidence for the NRG1 to HSCR and raised up a new susceptibility gene AUTS2 to a specific symptom for the disease.     

FEZ1/LZTS1 protein expression in ovarian cancer

The FEZ1/LZTS1 (FEZ1) gene maps to chromosome 8p22 and is frequently altered in human cancer. FEZ1 has been proposed as a candidate tumour suppressor gene and its loss may contribute to tumour progression. We have analysed the expression of FEZ1 protein in tissues from ovarian carcinomas in relation to clinico‐pathological variables, response to chemotherapy and disease‐free and overall survival. FEZ1 status was assessed by immunohistochemistry. Cytoplasmic staining for FEZ1 protein was absent or drastically reduced in 38% of tumours. FEZ1 protein expression was not related to tumour grade, histotype, disease‐free survival, or overall survival. On the contrary, it was significantly correlated with age and with FIGO stage of disease. This finding indicates that FEZ1 is involved in ovarian carcinogenesis. Moreover, loss of FEZ1 protein significantly predicted a complete treatment response in patients who received taxane‐based chemotherapy. In conclusion, the reduction or loss of FEZ1 protein could be an aid to the clinical management of patients affected by ovarian carcinoma. J. Cell. Physiol. 222: 382–386, 2010. © 2009 Wiley‐Liss, Inc.     

Transfer and expression of the human multiple drug resistance gene as potential human gene therapy

The human multiple drug resistance (MDR) gene has been used as a model for human gene transfer which could lead to human gene therapy. MDR is a transmembrane protein which pumps a number of toxic substances out of cells including several drugs used in cancer chemotherapy. Normal bone marrow cells express low levels of MDR and are particularly sensitive to the toxic effects of these drugs. There are two general applications of MDR gene therapy: (1) to provide drug-resistance to the marrow of cancer patients receiving chemotherapy, and (2) as a selectable marker which when co-transferred with a non-selectable gene such as the human beta globin gene can be used to enrich the marrow for cells containing both genes. We demonstrate efficient transfer and expression of the human MDR gene in a retroviral vector into live mice and human marrow cells including CD34⁺ cells isolated from marrow and containing the bulk of human hematopoietic progenitors. MDR gene transduction corrects the sensitivity of CD34⁺ cells to taxol, an MDR drug substrate, and enriches the marrow for MDR-transduced cells. The MDR gene-containing retroviral supernatant used has been shown to be safe and free of replication-competent retrovirus. Because of the safety of the MDR retroviral supernatant, and efficient gene transfer into mouse and human marrow cells, a phase 1 clinical protocol for MDR gene transfer into cancer patients has been approved to evaluate MDR gene transfer and expression in human marrow.     

急性髓系白血病ERG、MDR1及BAALC基因的表达水平及临床意义

目的:探讨急性髓系白血病(acutemyeloidleukemia,AML)患者骨髓单个核细胞的ETS相关基因(ETS related gene,ERG)、多药耐药基因1(Multidrug resistance gene 1,MDR1)、脑和急性白血病胞质(Brain and acute leukemia cytoplasmic,BAALC)基因的表达水平及临床意义。方法:选取90例成人AML患者为研究对象,均接受蒽环类药物诱导化疗联合阿糖胞苷等进行初步治疗,采用实时荧光定量检测PCR技术检测治疗后的骨髓单个核细胞ERG、MDR1、BAALC基因表达,并分析其与患者临床特征、危险度分层、治疗疗效、生存率的关系。结果:AML患者ERG、MDR1、BAALC基因均有强表达,三因子表达强弱同白细胞、血小板等临床指标无明显相关性(P0.05),不同危险度分层对应的ERG、MDR1、BAALC表达之间有明显差异,中危组、高危组患者表达强度均高于低危组(P0.05)。ERG、MDR1低表达组对应的疗效CR(93.1%%,89.6%)、OS(56.5%, 66.7%)较高表达组CR(61.4%, 81.3%)、OS(56.5%,66.7%)值更高(P0.05),不同BAALC表达者疗效CR及生存率OS比较差异无统计学意义(P0.05)。结论:AML患者单个核细胞的ERG、MDR1基因表达水平与其危险度分层、疗效和生存率呈负相关,以REG和MDR1同AML关系敏感,BAALC敏感度较低,联合检测REG、MDR1、BAALC基因表达可能提高AML患者危险度分层、疗效及预后判读的准确度。     

Induction of P‐glycoprotein and Bcrp at the rat blood–brain barrier following a subchronic morphine treatment is mediated through NMDA/COX‐2 activation

Subchronic morphine treatment induces P‐glycoprotein (P‐gp) up‐regulation at the blood–brain barrier. This study investigates the rate and extent to which P‐gp and breast cancer‐resistance protein (Bcrp) increase at the rat blood–brain barrier following subchronic morphine treatment. Rats were given increasing doses of morphine (10–40 mg/kg) or saline i.p. twice daily for 5 days. The brain cortex large vessels and microvessels were then mechanical isolated 6, 9, 12, 24, and 36 h after the last injection. The gene and protein expression of P‐gp and Bcrp in morphine‐treated and control rats were compared by qRT‐PCR and western blotting. The levels of Mdr1a and Bcrp mRNAs were not significantly modified 6 h post morphine, but the Mdr1a mRNA increased 1.4‐fold and Bcrp mRNA 2.4‐fold at 24 h. P‐gp and Bcrp protein expression in brain microvessels was unchanged 6 h post morphine and increased 1.5‐fold at 24 h. This effect was more pronounced in large vessels than in microvessels. However, extracellular morphine concentrations of 0.01–10 μM did not modify the expressions of the MDR1 and BCRP genes in hCMEC/D3 human endothelial brain cells in vitro. MK‐801 (NMDA antagonist) and meloxicam (cyclo‐oxygenase‐2 inhibitor) given after morphine treatment completely blocked P‐gp and Bcrp up‐regulation. Interestingly, misoprostol and iloprost, two well‐known agonists of prostaglandin E2 receptors induced both MDR1 and BCRP mRNA levels in hCMEC/D3. Thus, morphine does not directly stimulate P‐gp and Bcrp expression by the brain endothelium, but glutamate released during morphine withdrawal may do so by activating the NMDA/cyclo‐oxygenase‐2 cascade.     

Association of multiple drug resistance-1 gene polymorphism with multiple drug resistance in breast cancer patients from an ethnic Saudi Arabian population

Chemotherapy has been used widely to treat cancer, both as a systemic therapy and as a local treatment. Unfortunately, many types of cancer are still refractory to chemotherapy. The mechanisms of anticancer drug resistance have been extensively explored but have not been fully characterized. This study analyzed the occurrences of polymorphism (SNP) in the MDR1 gene in breast cancer patients and determined a possible association with chemotherapy. The study group included one hundred breast carcinoma patients who subsequently received chemotherapy (the regimen generally consisted of commonly used drugs such as cyclophosphamide, adriamycin, 5-fluorouracil, docetaxel and their combinations). Blood samples from 100 healthy individuals are used, as controls were also genotyped for the MDR1 gene. This investigation revealed a significant correlation with response to various regimens of chemotherapy showing a low response to therapy with the CT/TT genotype at (exon 12) 1236 codon (p < 0.001). These findings demonstrate, for the first time, that the polymorphisms in (exon 12) 1236 codon of the MDR1 gene greatly influence the drug response in patients from the Arab population of Saudi Arabia.     

The genetic component of human longevity: New insights from the analysis of pathway‐based SNP‐SNP interactions

In human longevity studies, single nucleotide polymorphism (SNP) analysis identified a large number of genetic variants with small effects, yet not easily replicable in different populations. New insights may come from the combined analysis of different SNPs, especially when grouped by metabolic pathway. We applied this approach to study the joint effect on longevity of SNPs belonging to three candidate pathways, the insulin/insulin‐like growth factor signalling (IIS), DNA repair and pro/antioxidant. We analysed data from 1,058 tagging SNPs in 140 genes, collected in 1825 subjects (1,089 unrelated nonagenarians from the Danish 1905 Birth Cohort Study and 736 Danish controls aged 46–55 years) for evaluating synergic interactions by SNPsyn. Synergies were further tested by the multidimensional reduction (MDR) approach, both intra‐ and interpathways. The best combinations (FDR<0.0001) resulted those encompassing IGF1R‐rs12437963 and PTPN1‐rs6067484, TP53‐rs2078486 and ERCC2‐rs50871, TXNRD1‐rs17202060 and TP53‐rs2078486, the latter two supporting a central role of TP53 in mediating the concerted activation of the DNA repair and pro‐antioxidant pathways in human longevity. Results were consistently replicated with both approaches, as well as a significant effect on longevity was found for the GHSR gene, which also interacts with partners belonging to both IIS and DNA repair pathways (PAPPA, PTPN1, PARK7, MRE11A). The combination GHSR‐MREA11, positively associated with longevity by MDR, was further found influencing longitudinal survival in nonagenarian females (p = .026). Results here presented highlight the validity of SNP‐SNP interactions analyses for investigating the genetics of human longevity, confirming previously identified markers but also pointing to novel genes as central nodes of additional networks involved in human longevity.